The three microelectrode voltage clamp technique will be used to investigate membrane charge movement (gating current) in frog skeletal muscle fibers. The experimental solution will be hypertonic and contain tetrodotoxin and tetraethylammonium to block mechanical activation and voltage dependent ionic currents. The metallochromic indicator dye arsenazo III will be injected iontophoretically through the current-passing electrode. The intracellular free (Ca2 ion ) transient will be determined by a differential measurement of the absorbance change of the Ca-dye complex. A 40 micron diameter measuring spot of light from a tungsten-halogen lamp will be focused on the muscle fiber at the point of voltage control. Changes in light transmittance will be measured at three wavelengths using a compound microscope, beam splitters and interference filters. A microspectrophotometer with the required sensitivity has been constructed. The objective of the research is to test the hypothesis that membrane charge movement in muscle is a gating current for Ca2 ion release from the sarcoplasmic reticulum. The hypothesis will be tested by observing whether charge movement and the Ca2 ion transient behave in a parallel way during 1) Prolonged depolarization, 2) Recovery from prolonged depolarization, 3) Exposure to various drugs and 4) Stretch to sacromere lengths from 2.0 to 3.6 micron. If the hypothesis proves tenable, subsequent work will attempt to establish the quantitative relationship between charge movement and Ca release. In addition to the principal line of investigation experiments will be conducted to verify the method and calibrate the optical signal.